title: Protein kinase inhibitors substantially improve the physical detection of T-cells with peptide-MHC tetramers.
abstract: Flow cytometry with fluorochrome-conjugated peptide-major histocompatibility complex (pMHC) tetramers has transformed the study of antigen-specific T-cells by enabling their visualization, enumeration, phenotypic characterization and isolation from ex vivo samples. Here, we demonstrate that the reversible protein kinase inhibitor (PKI) dasatinib improves the staining intensity of human (CD8+ and CD4+) and murine T-cells without concomitant increases in background staining. Dasatinib enhances the capture of cognate pMHC tetramers from solution and produces higher intensity staining at lower pMHC concentrations. Furthermore, dasatinib reduces pMHC tetramer-induced cell death and substantially lowers the T-cell receptor (TCR)/pMHC interaction affinity threshold required for cell staining. Accordingly, dasatinib permits the identification of T-cells with very low affinity TCR/pMHC interactions, such as those that typically predominate in tumour-specific responses and autoimmune conditions that are not amenable to detection by current technology.
authors.affiliation: Cardiff University School of Medicine, Heath Park, Cardiff, CF14 4XN, UK.
authors.forename: Anna
authors.initials: A
authors.lastname: Lissina
authors.forename: Kristin
authors.initials: K
authors.lastname: Ladell
authors.forename: Ania
authors.initials: A
authors.lastname: Skowera
authors.forename: Matthew
authors.initials: M
authors.lastname: Clement
authors.forename: Emily
authors.initials: E
authors.lastname: Edwards
authors.forename: Ruth
authors.initials: R
authors.lastname: Seggewiss
authors.forename: Hugo A
authors.initials: HA
authors.lastname: van den Berg
authors.forename: Emma
authors.initials: E
authors.lastname: Gostick
authors.forename: Kathleen
authors.initials: K
authors.lastname: Gallagher
authors.forename: Emma
authors.initials: E
authors.lastname: Jones
authors.forename: J Joseph
authors.initials: JJ
authors.lastname: Melenhorst
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Godkin
authors.forename: Mark
authors.initials: M
authors.lastname: Peakman
authors.forename: David A
authors.initials: DA
authors.lastname: Price
authors.forename: Andrew K
authors.initials: AK
authors.lastname: Sewell
authors.forename: Linda
authors.initials: L
authors.lastname: Wooldridge
mesh_terms: D000328:Adult; D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D000069439:Dasatinib; D018984:Epitopes, T-Lymphocyte / Q000276:immunology*; D005434:Flow Cytometry / Q000379:methods*; D015789:HLA-A2 Antigen / Q000276:immunology*; D006684:HLA-DR Antigens / Q000276:immunology; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D018448:Models, Immunological; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011743:Pyrimidines / Q000494:pharmacology*; D013194:Staining and Labeling / Q000379:methods; D013844:Thiazoles / Q000494:pharmacology*
chemical_list: D018984:Epitopes, T-Lymphocyte; D015789:HLA-A2 Antigen; D006684:HLA-DR Antigens; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D013844:Thiazoles; D000069439:Dasatinib